|
| Status |
Public on Aug 06, 2015 |
| Title |
Wenmai6-4 d Replicate 2 |
| Sample type |
RNA |
| |
|
| Source name |
Wenmai6 leaf sheath_4days
|
| Organism |
Triticum aestivum |
| Characteristics |
cultivar: Wenmai 6
phenotype: susceptible
tissue: Leaf sheath
age: day 30
|
| Treatment protocol |
Wheat were inoculated with small toothpicks harboring the well-developed mycelia of Rhizoctonia cerealis for 4 d,21d.
|
| Growth protocol |
All wheat plants were grown in a glasshouse at 22°C-14 h light (intensity of 300 μmol m-2 s-1) and 12°C-10 h dark conditions.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Trizol following manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
Cyanine-3/5 (Cy3/Cy5) labeled cRNA was prepared from 0.2 ug RNA using the Two-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
0.8 ug of Cy3/Cy5-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers' instructions. On completion of the fragmentation reaction, 55ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent-022297 Wheat Gene Expression Microarray (Agilent) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
Gene expression after Rhizoctonia cerealis infection for 4 d
S2-4
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities as the raw data. Raw data were normalized in quantile algorithm with Genespring 12.0(Agilent). Probe that at least 1 out of 2 samples flagged as Detected were maintained.
|
| |
|
| Submission date |
May 26, 2015 |
| Last update date |
Aug 06, 2015 |
| Contact name |
Xiuliang Zhu |
| E-mail(s) |
zhuxiuliang1987@126.com
|
| Organization name |
The Chinese Academy of Agricultural Sciences
|
| Street address |
Zhongguancun street 12#
|
| City |
Beijing |
| ZIP/Postal code |
100081 |
| Country |
China |
| |
|
| Platform ID |
GPL13636 |
| Series (1) |
| GSE69245 |
Wheat: Control vs. Pathogen-infected |
|
